Author ORCID Identifier

https://orcid.org/0000-0001-7230-2129

Date Available

7-20-2025

Year of Publication

2025

Document Type

Doctoral Dissertation

Degree Name

Doctor of Philosophy (PhD)

College

Medicine

Department/School/Program

Neuroscience

Faculty

Warren Joseph Alilain

Abstract

Efforts to improve treatment of spinal cord injury (SCI) often use animals to develop effective therapeutics, intending to improve the survival and the quality of life of persons with SCI. Over half of all SCIs in the United States occur at the cervical level and often cause locomotor deficits and life-threatening breathing dysfunction. Thus, my dissertation is focused on this important patient population.

After beginning with a description of translational research and surveying SCI from a clinical standpoint, I turned to describe experimental models of cervical SCI and more detailed neuroimmunological findings relevant to my work. Then, I conducted a literature review to compare and contrast animal care and use protocols for experimental SCI and clinical standards of care. I undertook this project to identify specific, practical areas in which veterinary treatments administered to animals used to study SCI could be amended to better reflect clinical care, thus improving their translational relevance. Findings demonstrated many key discrepancies which might contribute to why some promising pre-clinical discoveries fail to translate successfully to the clinic. Importantly, I found that animal subjects are often administered non-steroidal anti-inflammatory drugs (NSAIDs) following cervical-level injury, but NSAIDs are contraindicated following traumatic SCI in humans.

Clinical practices and treatments used in non-SCI contexts may serve as valuable candidates for reverse translation, enabling the evaluation of their off-label effects in preclinical experiments focused on functional recovery following SCI and potentially accelerating safe repurposing if efficacy is demonstrated. Bisphosphonate drugs like clodronate are already in clinical use for indications like osteoporosis, and liposomally encapsulated clodronate has shown efficacy in depleting hematogenous monocyte populations which differentiate into monocyte-derived macrophages (MDMs) within the spinal cord acutely after experimentally induced thoracic SCI, ameliorating tissue damage and improving locomotor recovery. Thus, we hypothesized that clodronate treatment would improve recovery of breathing and locomotor function following a C2 hemisection (C2Hx) model of cervical SCI in rats. During my dissertation work, changes to animal care and use guidelines serendipitously led to the inclusion of carprofen, an NSAID, in our study. This unexpected change prompted us to experimentally test how differences in the standard of care between our animal model and clinical practice might influence the effects of our repurposed therapeutic, liposomal clodronate.

Specifically, we treated adult rats intravenously with either liposomal clodronate or saline via the tail vein at days 1, 3, and 6 post-C2Hx. Carprofen treatment was administered subcutaneously immediately following SCI and twice more on days 1 and 2 post-injury. We used whole-body plethysmography to measure ventilatory function and the semi-automated CatWalk® gait analysis system to assess locomotor function in animals assigned to four separate treatment groups (clodronate, clodronate + carprofen, saline, and saline + carprofen) for up to 4 weeks post-SCI. Contrary to my initial hypothesis, I found that both liposomally encapsulated clodronate and carprofen impaired ventilatory recovery following C2Hx. However, in alignment with my hypothesis and consistent with prior findings in thoracic SCI, clodronate improved locomotor function on the side contralateral to injury. To reconcile these seemingly conflicting outcomes, I propose that clodronate-induced macrophage depletion may exacerbate lung inflammation and alter peripheral-to-central modulation of respiratory output—highlighting that the effects of these treatments may be specific to injury level and target organ system.

While neither clodronate and carprofen themselves are currently used in clinical practice, their respective drug classes—bisphosphonates and non-steroidal anti-inflammatory drugs—are widely employed in human medicine for other indications. As such, this work serves as an example of reverse translation: bringing clinically utilized therapeutics back into preclinical experimentation to evaluate their potential for off-label use in SCI. Coupled with efforts to improve the clinical relevance to preclinical models of SCI, the work described within this dissertation ultimately serves to advance efforts to improve care for individuals living with spinal cord injury.

Digital Object Identifier (DOI)

https://doi.org/10.13023/etd.2025.268

Funding Information

This work was financially supported by supported by the National Institutes of Health (R01NS116068 to John Gensel (JCG) and Warren Alilain (WJA) from 2021-2025; R21NS137256 to Meifan Chen (MC) and WJA from 2024-2025; T32AA027488 to directors Mark Fillmore and Mark Prendergast, intramurally awarded to Aaron Silverstein from 2022-2024) and a Pilot Grant from the Craig H. Neilsen Foundation (to MC and WJA from 2024-2025).

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